Methyl-CpG binding proteins (MBPs) are central players of epigenetic regulation by translating DNA methylation into phenotypic changes. Mutations in MeCP2 result in Rett Syndrome and certain autism- spectrum disorders. Alterations in MBD1 are found in a subset of autistic patients and intellectual disabilities. We have demonstrated that MBD1 null mice display learning impairment and autism-like behavioral deficits. Thus, both MeCP2 and MBD1 are autism-associated genes (SFARI). We have shown that both MeCP2 and MBD1 are expressed in immature stem cells and new neurons in the postnatal brains and regulate neurogenesis and neuronal maturation. We found that such regulations are at least in part through controlling the expression of noncoding microRNAs. Although many studies have focused on MeCP2 target identification, MeCP2 targets in specific neural cell types remain largely unknown. In addition, little has been done to identify the targets of MBD1 in the brain. Literatures have suggested that MBD1 and MeCP2 may have different sets of targets, however both protein exhibit similar heterochromatin binding and share at least several noncoding gene targets. Functional relationship between these two proteins during neuronal development has not been explored. We observed that mice lacking both MBD1 and MeCP2 die much earlier than single mutant mice, at postnatal day 21, which coincides with the postnatal synaptogenesis and neuronal maturation period as well as the dynamic changes in DNA methylation in neurons. Therefore a lack of both MBD1 and MeCP2 during this period is extremely detrimental but the reason is unknown. In addition, despite the apparent functional compensations between these two proteins, questions remained why MBD1 cannot fully compensate for MeCP2 deficiency and which genomic binding targets might be responsible for this lack of compensation. A direct comparison of genome-wide MBD1 and MeCP2 targets will yield valuable information for understanding MBD regulation of neurodevelopment and disorders. The PI and collaborators are uniquely suited to fill these knowledge gaps. In this exploratory R21 proposal, we aim to take advantage of a MeCP2-FLAG line created by collaborator Dr Chang and our newly created MBD1-FLAG mouse line to directly compare the targets between MBD1 and MeCP2 in developing brain. We aim to test the hypothesis that MBD1 and MeCP2 have both common and unique targets which underlie their specific functions during postnatal development. We will first identify genomic binding profiles of these proteins. We will then determine whether the genomic binding specificities have impact on gene expression and neuronal maturation. The proposed study is built upon the complimentary expertise of the PI and collaborators. The results of this study will provide much needed understanding in biological functions of MBP and its significance in gene regulation. Since both MBPs are important for autism and neurodevelopmental disorders, the study will unveil the novel genes and pathways involved in developmental disorders.